Enkephalin and dynorphin are opioid peptides found in various brain regions including the striatum. The striatal opioid neurons express dopamine receptors and are regulated by dopaminergic input. We have found previously that chronic treatment of pregnant monkeys with cocaine causes increased expression of the opioid peptides in the rostral forebrain of their fetuses. Enkephalin binds to both mu- and delta-opioid receptors to inhibit adenylyl cyclase and cellular activity. The present studies were performed to determine the ontogeny of mu-opioid receptors and to explore if gestational cocaine exposure would decrease the expression of mu receptors in the fetal monkey brain. A fragment of the monkey mu-receptor gene was cloned using reverse transcription and polymerase chain reaction (PCR). The resulting 293 bp PCR-product was subcloned, and used in nuclease protection assay and in situ hybridization. Mu receptor mRNA was detected in the thalamus of day 40 fetuses. By day 60 faint expression of mu receptor mRNA was found in the striatum, olfactory bulbs, the globus pallidus, hypothalamus and brain stem. The highest levels of mu receptor mRNA were observed in the dorsomedial and reticular nuclei of the thalamus and the medial habenula. Although, the fetal brain on day 70 was much more developed than on day 60, the overall distribution of mu receptor mRNA was similar to that found on day 60. In contrast, the cellular concentration of mu receptor mRNA had increased in most brain regions. The effects of fetal cocaine exposure was explored in dissected tissues from day 70 fetuses using nuclease protection assay analysis. This quantitative analysis confirmed the data obtained from the in situ hybridization experiments. In addition, we found that fetal cocaine treatment caused a significant decrease in mu-opioid receptor mRNA concentration in the diencephalon. These data together with our previous results suggest that prolonged gestational cocaine exposure has rather profound effects on the endogenous opioid systems in the fetal brain.